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Structural basis for the initiation of eukaryotic transcription-coupled DNA repair

Xu, Jun and Lahiri, Indrajit and Wang, Wei and Wier, Adam and Cianfrocco, Michael A. and Chong, Jenny and Hare, Alissa A. and Dervan, Peter B. and DiMaio, Frank and Leschziner, Andres E. and Wang, Dong (2017) Structural basis for the initiation of eukaryotic transcription-coupled DNA repair. Nature, 551 (7682). pp. 653-657. ISSN 0028-0836. PMCID PMC5907806.

[img] PDF - Accepted Version
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[img] PDF (Supplementary Figure 1, the gel source data and Supplementary Tables 1-2) - Supplemental Material
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[img] PDF (Life Sciences Reporting Summary) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 1 : Sequence alignment of the ATPase core domains of CSB family members) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 2 : Cryo-EM reconstructions of the Pol II–Rad26 and Pol II EC complexes) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 3 : Validation of Rosetta models for the Pol II–Rad26 complex and Pol II EC) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 4 : Cryo-EM reconstruction of a Pol II EC containing a CPD lesion) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 5 : Alignment of the HD2-1 region of CSB and non-CSB members of the Swi2/Snf2 superfamily of ATPases) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 6 : The strength of base pairing at the upstream fork of the transcription bubble, not CPD lesions at downstream fork, affects the interaction of Rad26 with Pol II EC) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 7 : Overlap between the binding sites of Rad26 and Spt4–Spt5 on Pol II) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 8 : Alignment between Snf2 and Rad26) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 9 : Unified model for three-step DNA lesion recognition and verification for both TCR and GG-NER) - Supplemental Material
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[img] Image (JPEG) (Extended Data Table 1: Specific P values) - Supplemental Material
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Eukaryotic transcription-coupled repair (TCR) is an important and well-conserved sub-pathway of nucleotide excision repair that preferentially removes DNA lesions from the template strand that block translocation of RNA polymerase II (Pol II). Cockayne syndrome group B (CSB, also known as ERCC6) protein in humans (or its yeast orthologues, Rad26 in Saccharomyces cerevisiae and Rhp26 in Schizosaccharomyces pombe) is among the first proteins to be recruited to the lesion-arrested Pol II during the initiation of eukaryotic TCR. Mutations in CSB are associated with the autosomal-recessive neurological disorder Cockayne syndrome, which is characterized by progeriod features, growth failure and photosensitivity1. The molecular mechanism of eukaryotic TCR initiation remains unclear, with several long-standing unanswered questions. How cells distinguish DNA lesion-arrested Pol II from other forms of arrested Pol II, the role of CSB in TCR initiation, and how CSB interacts with the arrested Pol II complex are all unknown. The lack of structures of CSB or the Pol II–CSB complex has hindered our ability to address these questions. Here we report the structure of the S. cerevisiae Pol II–Rad26 complex solved by cryo-electron microscopy. The structure reveals that Rad26 binds to the DNA upstream of Pol II, where it markedly alters its path. Our structural and functional data suggest that the conserved Swi2/Snf2-family core ATPase domain promotes the forward movement of Pol II, and elucidate key roles for Rad26 in both TCR and transcription elongation.

Item Type:Article
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URLURL TypeDescription ReadCube access CentralArticle
Wang, Wei0000-0002-5257-7675
Dervan, Peter B.0000-0001-8852-7306
Additional Information:© 2017 Macmillan Publishers Limited, part of Springer Nature. Received: 21 May 2017; Accepted: 18 October 2017; Published online: 22 November 2017. We thank the Wang and Leschziner laboratories for discussions. D.W., A.E.L. and P.B.D. were supported by National Institutes of Health (NIH) grants GM102362, GM102362-S1 (D.W.), GM092895 (A.E.L.), and GM27681 (P.B.D.). M.A.C. acknowledges support from the Damon Runyon Cancer Research Foundation. We thank the UCSD cryo-EM Facility, where all data was collected. We used the Extreme Science and Engineering Discovery Environment (XSEDE) for computing allocations (MCB160121 to D.W.), supported by NSF grant ACI-1548562. Author Contributions: J.X. prepared the proteins with help from W.W. and J.C. and performed the biochemical analyses. A.H. and P.D.B. provided the Py-Im chemical agent. I. L. collected the EM data with help from A.W. I.L. performed data processing and refinement with help from M.A.C. I.L. and F.D. generated the atomic models with homology models generated by J.X., W.W. and D.W. D.W. and A.E.L. wrote the manuscript with help from all laboratory members. D.W. and A.E.L. directed and supervised the research. The authors declare no competing financial interests.
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Damon Runyon Cancer Research FoundationUNSPECIFIED
Issue or Number:7682
PubMed Central ID:PMC5907806
Record Number:CaltechAUTHORS:20171127-145158492
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Official Citation:Xu, J., Lahiri, I., Wang, W. et al. Structural basis for the initiation of eukaryotic transcription-coupled DNA repair. Nature 551, 653–657 (2017).
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:83462
Deposited By: Tony Diaz
Deposited On:27 Nov 2017 23:29
Last Modified:15 Apr 2020 22:03

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